A loop-mediated isothermal amplification assay combined with lateral flow dipstick for rapid detection of Aphelenchoides besseyi
Jiue-in Yang: National Taiwan University
<div>The rice leaf and bud nematode, <em>Aphelenchoides besseyi,</em> is an economically important plant parasitic nematode and a concerned quarantine pest to many countries globally. It causes the white tip disease on rice, and some susceptible rice varieties were recorded with 70% yield loss. Early detection of the <em>A. besseyi</em>-infected rice seedling during the nursery stage could prevent the spread of this seed-borne pathogen. A loop-mediated isothermal amplification (LAMP) assay combined with lateral flow dipstick was developed for the rapid screening and early detection purposes. A set of 4 primers, named ABID37, was designed to target the mitochondria cytochrome oxidase I (COI) gene. The ABID37 LAMP assay was optimized to complete within 60 minutes at 63°C, with the sensitivity of 10<sup>4 </sup>copies cloned DNA plasmid per µl. The assay is highly specific and can effectively exclude detection of the very closely related fern biotype of <em>A. besseyi</em> and other common aboveground plant-parasitic nematodes. The specific lateral flow dipstick was successfully developed for immediate ABID37 LAMP result analysis without agarose gel electrophoresis, and no false positive was detected when examined with mix total DNA of rice and <em>A. besseyi</em>.</div>
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